Non-melanoma skin cancer (NMSC) is now the most common form of cancer in white populations and incidence rates for NMSC and malignant melanoma are increasing steadily world wide. While ultraviolet radiation (UVR) is the greatest environmental risk factor for skin cancer, skin pigmentation phenotype appears to be the most important genetic determinant of risk.
The melanocortins include a family of peptide hormones that induce pigmentation by interaction with melanocortin 1 receptors (MC1R) in the epidermis (see Hadley, M E. The melanotropic hormones, In: Brake, D., editor. Endocrinology, 4th Edition, Simon & Schuster; 1982; pp. 153-76). The primary pigmentary hormone that is released from the pars intermedia of the pituitary gland in some non-human animals, and from UV-B exposed keratinocytes in human skin, is α-melanocyte stimulating hormone (α-MSH). This 13 amino acid peptide binds to MC1R to induce cyclic AMP-mediated signal transduction leading to the synthesis of melanin polymers from DOPA precursors. Two types of melanins can be expressed in humans. The brownish-black pigment eumelanin is believed to convey protection from sun damage, whereas the reddish, sulfur-containing pigment, phaeomelanin is often expressed in light-skinned human populations that report a poor tanning response to sunlight. These poorly-tanning, easily-burning populations, often possess defects in the MC1R gene [28], and are generally thought to be at a greater risk of developing both melanoma and non-melanoma skin cancers [5, 21].
α-MSH binds MC1R to stimulate both eumelanogenesis, by upregulating tyrosinase activity, and melanocyte proliferation, through activation of adenylate cyclase [1, 6, 13]. Eumelanin is known to have photoprotective properties as it is resistant to photodegradation and has the ability to quench reactive oxygen radicals [19, 20]. Studies investigating whether variant alleles affect the interaction between α-MSH, MC1R and the downstream processes have found that the MC1R gene is highly polymorphic and variants such as Arg151Cys, Arg160Trp and Asp294His, are associated with fair skin colour and red hair, characterised by a low melanin content and a low eumelanin to phaeomelanin ratio [5, 16, 28]. Several variants, including the above-mentioned, have since been associated with an increased risk of skin cancer independent of pigmentation phenotype [4, 17, 21]. It has also been demonstrated that the variants Arg142His, Arg151Cys, Arg160Trp and Asp294His are loss-of-function alleles, and one effect of having these variant alleles is a decrease in the binding affinity of MC1R to α-MSH [11, 15, 22, 24, 27]. Investigations conducted into human MC1R variants have established that either “loss-of-function” or “diminished function” mutations in the MC1R gene sensitise human melanocytes to the DNA damaging effects of UV radiation, which may increase skin cancer [12, 24, 25].
Although (α-MSH stimulates natural skin protection, the process requires harmful UVR. It has previously been disclosed that a super-potent derivative of α-MSH, Melanotan-1, (Nle4-D-Phe7-α-MSH), can induce tanning in human volunteers [18]. Melanotan (MT-1), contains two amino acid substitutions and exhibits a 10- to 100-fold increased activity in frog and lizard bioassays for pigmentation [25], increases melanogenesis and tyrosinase activity in human melanocytes, and more specifically, induces significant increases in the eumelanin content of melanocytes while having a lesser effect on the levels of phaeomelanin [7, 13, 14]. Several studies have assessed the pharmacokinetic and tanning effects of Melanotan in humans and found a significant increase in eumelanin, but not phaeomelanin, content in skin [7, 18]. Although melanotropins have been postulated to effect immunologic changes, all of the prior trials reported only minimal side effects such as facial flushing and transient GI upset, unless doses greater than those needed for tanning were administered.
U.S. Pat. No. 4,457,864 (issued Jul. 3, 1984), discloses analogues of α-MSH, including Nle4-D-Phe7-α-MSH. Cyclic analogues of α-MSH are disclosed in U.S. Pat. No. 4,485,039 (issued Nov. 27, 1984). The use of these and other analogues of α-MSH for stimulating the production of melanin by integumental melanocytes is disclosed in Australian Patent No. 597630 (dated Jan. 23, 1987) and U.S. Pat. No. 4,866,038 (issued Sep. 12, 1989), U.S. Pat. No. 4,918,055 (issued Apr. 17, 1990) and U.S. Pat. No. 5,049,547 (issued Sep. 17, 1991). Australian Patent No. 618733 (dated May 20, 1988), and U.S. Pat. No. 5,674,839 (issued Oct. 7, 1997), U.S. Pat. No. 5,683,981 (issued Nov. 4, 1997) and U.S. Pat. No. 5,714,576 (issued Feb. 3, 1998) disclose further linear and cyclic α-MSH fragment analogues, and the use of these biologically-active analogues in stimulating melanocytes. The contents of all these published Australian and US patents are incorporated herein by reference.
In work leading to the present invention, it has been demonstrated that notwithstanding the significantly reduced response to α-MSH of human melanocytes having either “loss-of-function” or “diminished function” mutations, Melanotan is effective in inducing melanogenesis in human subjects having MC1R variant alleles. In particular, it has been demonstrated that significant increases in melanin density can be induced in such subjects by use of Melanotan, in some cases leading to melanin density levels similar to the levels in subjects having the wild-type MC1R.
Accordingly, the method of the present invention enables the induction of melanogenesis in human subjects having a “loss-of-function” or “diminished function” mutation(s) in the MC1R gene, leading to increased melanin density levels in these subjects and reduced risk of skin cancer.